JPH0338824B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Application Field] This invention provides a method for producing a food composition containing 5 mg or more of inositol triphosphate (IP ₃ ) per 100 g of the composition, and a food composition containing such an amount of IP ₃ . Regarding. BACKGROUND OF THE INVENTION There is an increasing need to prevent many of the negative effects of urbanization, such as the environmental hazards caused by the introduction of hazardous materials such as heavy metals and irradiation. Additionally, the development of health foods and nutritional supplements has increased the need to prevent the harms of smoking and other bad habits. Already in 1900, several researchers discovered the organophosphorus compound phytic acid, namely 1,2,3,4,
reported the discovery in plants of 5,6-hexakis(dihydrogen phosphate) myo-inositol (also sometimes referred to as inositol pentaphosphate). The content of phytic acid in different plants varies considerably. The content in grains is usually about 0.5-2%, with some exceptions. A little bit of polished rice
Brown rice contains as much phytic acid as 2.2%, while it has a level of 0.1%. Beans account for approximately 0.4-2%;
Oil plants contain about 2-5% and pollen 0.3-2%. The content of phytic acid in plants changes during the growing period. This content is also influenced inter alia by the climate. The presence of inositol pentaphosphate (IP ₅ ) and inositol tetraphosphate (IP ₄ ) in a few plants has been reported in the literature. Furthermore, it is known that phosphoric acid derivatives lower than IP ₆ are found during grain germination. For example, the end products during germination are inositol and phosphate. The use of IP ₆ has been described in several scientific publications. Many of the authors of these papers have observed some negative effects when consuming IP ₆ or IP _6- containing products. For example, if dogs are bred with too high a content of IP ₆ , rickets will result, for example. In humans, zinc deficiency and consequent slow growth in children have been observed. Anemia was observed primarily in women. Because of the above negative effects on mineral balance in humans or animals, attempts have been made to minimize the uptake of IP ₆ and its derivatives. Cadmium has also been found to be harmful to human health. This mineral is generally present at low levels in our environment, and the amount of cadmium we are exposed to depends on several factors. The presence of cadmium and its availability in soil varies by region, and uptake into plants growing in areas with relatively low pH values is relatively high. Industrial activities, primarily metal handling, can release cadmium into the air, soil, and water. Cadmium in soil can be absorbed by plants and therefore enter the diet of humans and animals. The most important routes of exposure to cadmium are through smoking, food, and to some extent drinking water. Although only a small portion of cadmium in food is absorbed, cadmium is primarily absorbed in the intestines and through the lungs. Average cadmium intake through food is approximately
It is calculated to be 50 μg/day, but there are large variations between different geographical regions and between individuals. Data from smokers show that as much as 50% of inhaled cadmium can be absorbed. Some studies have
It shows that blood and organ levels of cadmium are twice as high in smokers compared to non-smokers.
The excretion of cadmium from the human body is slow and 10
A half-life of ~30 years has been reported. This means that
This means that cadmium accumulates in the body.
80-90, which is the main part of the accumulated cadmium
% binds to metallothionein, a protein found primarily in the liver and kidneys. The formation of metallothionein is induced by metals, primarily zinc and cadmium. The binding of cadmium to metallothionein is very strong and results in cadmium detoxification. The remaining cadmium is present in the body. That is, cadmium that is not bound to metallothionein is distributed in many organs of the body, with relatively low concentrations in the intestines, lungs (particularly the lungs of smokers), the circulatory system (heart, artery walls, spleen), and glands such as the pancreas and prostate. Distributed at high levels. Among the negative effects, cadmium improves the body's elastin/
It is known that it can affect the elastase system. In addition, cadmium affects several different enzymes in the body, such as Na ⁺ , K ⁺ (Mg ²⁺ )-ATP-ase and Ca ²⁺ , Mg ²⁺ -ATP-ase, which are important in ion transport systems. ) is known to have an impact on Other enzymes are cytochrome-P450-enzymes that hydrolyze steroids, fatty acids, aromatic compounds and toxic compounds. Other important enzymes inhibited by cadmium are glutathione-peroxidase and superoxide dismutase, which provide protection against the occurrence of peroxidation. Zinc dependent enzymes such as leucine aminopeptidase are also inhibited by cadmium. Results from numerous animal studies obtained over many years show negative effects even at very low concentrations of cadmium. This would mean that a large part of the population is negatively affected, and this is especially so for smokers. Epidemiological studies show an association between cancer, hypertension and cardiovascular diseases (eg arteriosclerosis, myocardial infarction, acute cardiac death) and the presence of cadmium in the environment. Exposure to cadmium is also thought to be a factor that increases the risk of geriatric diabetes. In addition, cadmium is harmful to the kidneys, lungs (fibrosis, emphysema, cancer), vascular walls (fat accumulation, arteriosclerosis, vascular wall contraction, elasticity, damage to the endothelium), prostacyclin production, prostate, heart (conduction system, contractile force), ),
It can have negative effects on the placenta, testicles, and central nervous system. Cadmium can also induce the production of free radicals and thereby cause lipid peroxidation, which is important in the development of other diseases such as rheumatism. Allergies and bronchitis may also be associated with cadmium exposure. Knowledge of the negative effects of cadmium on humans and animals has increased considerably over the decades. Aiming at countering the above-mentioned negative effects of heavy metals, such as cadmium, and in various ways the negative effects of free radicals generated by metals such as iron, aluminum and cadmium or by irradiation, very powerful treatments have been used for many years. A great deal of research effort has been made. Needless to say, the harms of smoking have also been studied for a long time. [Summary of the Invention] According to the present invention, the above-mentioned negative effects observed in humans and animals can be avoided or at least reduced by consumption of specific inositol triphosphates (IP ₃ ). It was hot. The invention therefore provides a method for producing a food composition, which composition has an _IP3 of 5 mg or more per 100 g of the composition.
Contains. It is generally preferred to use IP ₃ in salt form. However, it can also be used in the acid form if desired. [Specific Description] According to the present invention, a food composition having an IP ₃ content of 5 mg or more per 100 g of the composition has been achieved. Very often the IP ₃ content is 20 mg or more per 100 g of the composition. Advantageously, the content of IP ₃ in the food composition
5 to 500 mg per 100 g, preferably 20 to 500 mg, more preferably 50 to 500 mg, 100 to 500 mg, or 150 to
In the 500mg range. This composition can be used as an additive or intermediate concentrate to increase the IP ₃ content of other food products. In this case, the content of IP ₃ in the intermediate concentrate should be at least 20 mg/100 g of concentrate. Generally, however, the IP ₃ content in this concentrate is higher, advantageously between 50 mg and 100 g, and preferably 75 g, respectively, per 100 g of concentrate.
mg~80g, 100mg~80g, 150mg~60g, 200mg~
60g, 250mg to 50g, or 300mg to 50g. Preferably, the IP ₃ content of the concentrate should be as high as possible. Intermediate concentrates can be used in many different forms, such as powders, tablets, capsules and granules. However, it can also be used in liquid form, for example in the form of an aqueous solution. IP ₃ is preferably D-myo-inositol-
1.2.6-triphosphate, D-myo-inositol-1.2.5-triphosphate, myo-inositol-1.2.3-triphosphate, L-myo-
selected from the group consisting of inositol-1.3.4-triphosphate, and D-myo-inositol-1.4.5-triphosphate, and mixtures thereof. Among these isomers, D-myo-inositol-1.2.6-triphosphate is preferred. Often 20 to 100% by weight, preferably 40 to 100% by weight of IP ₃ consists of D-myo-inositol-1.2.6-triphosphate. According to a suitable method for producing IP ₃ , the material containing inositol hexaphosphate (IP ₆ ) is enzymatically degraded by phytase enzymes. This IP ₆ can be provided as a pure substance or in the form of an IP _6- containing source, such as wheat bran. Phytase enzymes can be found, for example, in plants, seeds, and microorganisms. Enzymatic treatment of IP ₆ results in hydrolysis of different lower inositol phosphates, namely inositol pentaphosphate (IP ₅ ), inositol tetraphosphate (IP ₄ ), inositol triphosphate (IP ₃ ), and inositol diphosphate. ate (IP ₂ ) and inositol monophosphate (IP ₁ )
A mixture of is formed. Generally, this hydrolysis is carried out at 20°C to 70°C and 4 to 8°C.
It is carried out at the PH of This hydrolysis is suitably stopped when about 30-60% of the total ester phosphorus is liberated. At this stage, a high proportion of the desired IP ₃ among isomers is formed by hydrolysis of the IP ₆ containing material. After this, the resulting mixture of inositol phosphates was separated by chromatography and IP ₃
The containing fraction can be isolated. Preferably,
This is done in a column. If the IP ₃ fraction contains more than one isomer, it is subsequently separated in another chromatographic separation step. IP ₃ can be obtained as a salt or as the acid itself. Salt forms are preferred because they are easier to obtain in pure and concentrated form than acids. The salt form of the IP ₃ isomer can be readily obtained from the acid form using standard methods. Thus, salts such as alkali metal salts and alkaline earth metal salts such as lithium, sodium, potassium, calcium or magnesium salts can be obtained. In addition, however, zinc salts are very useful, as are NH ₄ ⁺ salts and organic amine salts. Examples of amines include triethanolamine, diethanolamine, triisopropanolamine, N,N-diethyl-2-amino-2-methyl-1-propanol, N,N-dimethylethanolamine, tetrabutylamine and cyclohexylamine. Additionally, other salts could be used. Particularly preferred salts are physiologically acceptable salts. Preferably, the distribution curve showing the content of different inositol phosphates has a maximum for IP ₃ , and preferably only one maximum, which means that the IP ₃ content is greater than the IP ₂ and/or IP ₄ content. means. Generally, the proportion of IP ₃ is more than 10% of the total content of inositol phosphate. Sometimes the compositions contain, in addition to IP ₃ , IP ₄ and/or inositol diphosphate (IP ₂ ).
In this case, preferably at least 40% by weight of the total amount of inositol phosphate in the composition is IP ₃ ;
On the other hand, 30-85% by weight of the remaining inositol phosphates are IP ₂ and IP ₄ . In such compositions, IP ₃ is D-myo-inositol-1.2.6-
It can consist essentially of triphosphate.
However, it is also possible to use other IP ₃ isomers, especially those mentioned above, in such food products. For example, depending on the form of the composition, IP ₃ can be used in salt form or in acid form. The acid form is generally used as a liquid, preferably an aqueous solution. In the salt form IP ₃ as a dry product:
Alternatively, it can be used as a liquid, preferably as an aqueous solution. When IP ₃ is present as a salt, this salt is generally selected from the group mentioned above. The invention provides a method for producing a food composition, which food product is initially substantially free of _IP3 . The method comprises adding a source of IP ₃ to the composition in an amount sufficient to provide a concentration of IP ₃ of 5 mg or more per 100 g of the composition. The source of IP ₃ may be separately produced IP ₃ itself or it may be IP ₆ , IP ₅ and/or IP ₄ in the presence of phytase for enzymatic production of IP ₃ . "Initially substantially free of IP ₃ " means that a food composition manufactured according to conventional methods contains a substantial amount of IP 3.
Intended to mean does not contain IP ₃ . The content of IP ₃ will therefore be less than 3 mg, usually less than 2 mg, and most often less than 1 mg per 100 g of the composition. The invention also includes a method of making a food composition,
In this method, in the materials constituting the composition, a concentration of phetase and an inositol selected from the group consisting of inositol tetraphosphate ( _IP4 ), inositol pentaphosphate ( _IP5 ), and inositol hexaphosphate ( _IP6 ) are determined. Establish the content of phosphate and, in at least one stage of the manufacturing process, per 100 g of the composition.
20mg or more of inositol triphosphate ( _IP3 )
The processing time, temperature and pH are adjusted so that the incubation can be carried out so that the content of
Control the value. In the incubation step, IP ₆ ,
At least a portion of the inositol phosphate selected from IP ₅ and IP ₄ is purified by a phytase enzyme.
Enzymatically degraded to _IP3 . The proportion of the original inositol phosphate content that is converted to IP ₃ depends on the manufacturing parameters, e.g. incubation time,
It can be controlled within wide limits by varying the temperature and PH. If the plants and seeds contain inositol hexaphosphate, the phytase enzyme can be present in these. According to the invention, therefore, it is not necessary in all cases to add enzymes when using plant or seed products as starting materials. If the natural products mentioned have too low an enzymatic activity, or if IP ₆ , IP ₅ or IP ₄ or mixtures thereof are used as starting material, a phytase enzyme, for example derived from bran, can be added. Yeast can advantageously be used as a source of phytase. Preferably, baker's yeast is used. According to the invention, for example, Swedish baker's yeast produced by Ja¨stbolaget (Sweden) and Radiyameki, Finland & Hoef Abliken AG
(Rajama¨ki, Finland and Hefefabriken AG)
(Switzerland) was used. When using yeast according to this invention, it is very surprising that only one isomer, namely D-
It has been established that myo-inositol-1.2.6-triphosphate is obtained. Of course, if only this isomer is desired, the use of yeast is a very valuable method. Appropriate temperature during incubation, typically 20°C
Hydrolysis is carried out at ~70°C, preferably from 30°C to 60°C, and at a PH of 4-8. To stop hydrolysis at the desired level, the enzyme can be destroyed or inactivated, for example by rapid heating of the hydrolyzed starting material. The method of the invention can be modified in various ways, depending for example on the starting materials chosen.
The starting material can, for example: 1 have a certain amount of _IP6 , _IP5 and/or _IP4 . 2. Cannot contain IP ₆ , IP ₅ or IP ₄ . In the first example above, in the final food product
Different possibilities exist to achieve the desired content of IP ₃ . For example, the method described above of hydrolyzing inositol phosphates to _IP3 by phytase can be used. If the content of IP ₆ , IP ₅ and/or IP ₄ in the starting materials is not high enough, they can be added. In this way, the IP ₃ content in the final product can be increased. The above method can also be used for the production of intermediate products with the desired IP ₃ content, which can be added to the starting materials for food compositions. Intermediate products can also be introduced at later stages of food product manufacture. The method for producing IP ₃ and its isomers itself is described in a patent application filed on the same day as this application entitled "Inositol triphosphate and method for producing the same" filed by the same applicant as the present patent applicant. There is. In the second example above, where the starting material does not contain IP ₆ , IP ₅ or IP ₄ , these inositol phosphates can be added together with phytase if it is lacking. The hydrolysis method described above can then also be used to obtain the desired concentration of IP ₃ in the final product. Alternatively to the above method, the intermediate product described above may be
It can be added to the starting materials for the production of food compositions or at a later stage. In any of the above methods, a concentrated form of
IP ₃ can be added at a later or final stage of composition production. In another aspect of the invention, a method of making a food composition is provided, the composition first comprising:
containing less than about 10 mg of IP ₃ per 100 g, and in this method by adding IP ₃ or a source of IP 3 or by initially containing IP ₆ selected from the group consisting of IP ₆ , IP ₅ and IP ₄ . By converting inositol phosphate by enzymatic method,
The content of IP ₃ is increased to 20 mg or more per 100 g of the composition. In such a method of manufacturing a composition of the invention, said composition preferably comprises a cereal-based material, such as a breakfast food.
cereals), cakes, biscuits and bread. The composition may also be selected from the group consisting of confectionery, thiokolate and chewing gum. Often, very preferably, the composition is also a vegetable, a drink, a soup or a milk product, such as a yoghurt. The composition initially contains less than about 20 mg per 100 g of the composition.
In other embodiments of this invention containing IP ₃ ,
In a similar manner, the content of IP ₃ was determined to be 50% per 100g of the composition.
mg or more. composition first composition
In other embodiments of the invention containing about 50 mg of IP ₃ per 100 g, the content of IP ₃ is increased to 100 mg or more per 100 g of the composition by a similar method. The IP ₃ content in the composition can vary within wide limits. Preferably it has an IP ₃ content of 20 to 500 mg, such as 50 to 500 mg, 100 to 500 mg or 150 to 500 mg per 100 g of composition. Regarding bread foods,
This interval is advantageously 20 g per 100 g of dry bread food.
~500mg, preferably 100-500mg and most preferably 150-500mg or 200-500mg IP ₃ .
The daily intake of IP ₃ is at least 10mg, preferably 50mg
That's all. In the production of the liquid composition of this invention, IP ₃
The content of can also be varied within a wide range.
Generally, the content of IP ₃ is between 5 and 5 per 100 g of liquid composition.
500mg, such as 10-500mg, 20-500mg, or 5-500mg
It is 300mg. The liquid composition can be, for example, a drink or a soup. According to the invention, the concentration of IP ₃ is 100% in the dry composition.
Food compositions can be provided that have 20 mg/g or more. IP ₃ is produced by adding IP ₃ to the composition and/or adding phytase and an inositol phosphate selected from the group consisting of IP ₄ , IP ₅ and IP ₆ . Particular preference is given to bread products within the scope of this invention. That is, the concentration of IP ₃ per dry composition
20 mg or more and the degree of hydrolysis of naturally occurring IP ₆ and/or added IP ₆ is from 20 to
A bread food composition is provided which is 90%, preferably 40-70%. Therefore, the ratio of inorganic phosphorus to the total amount of phosphorus is 20% or more, preferably 40% or more. In certain bread food compositions, the initial content of IP ₃ is less than 150 mg per 100 g of the composition;
And the initial content of IP ₆ is 200 mg per 100 g of composition
That's all. And the content of IP ₃ in the final composition increases due to the decomposition of IP ₆ . The IP ₃ isomer described above has the following structure. D-myo-inositol-1.2.6-triphosphate (wherein X is hydrogen, at least one monovalent, divalent, or polyvalent cation, or a mixture thereof, n is the number of ions, and z is the charge of each ion.) D -Myo-inositol-1.2.5-triphosphate (In the formula, X, n and z have the above meanings.) Myo-inositol-1.2.3-triphosphate (wherein X, n and z have the above meanings) L-myo-inositol-1.3.4-triphosphate (wherein X, n and z have the above meanings) D-myo-inositol-1.4.5-triphosphate (wherein X, n and z have the meanings given above) In each of the above formulas, n ranges from 6 to 1 (inclusive) and z ranges from 1 to
6 (inclusive). Preferably, n is 3 to 6 inclusive and z is 3, 2, or 1. The compositions of this invention positively impact living organisms in a number of ways. However, this composition is primarily intended to avoid or alleviate conditions caused, induced or promoted by heavy metals, essentially cadmium, or diseases associated with these heavy metals. Furthermore, this composition is intended to provide good effects to smokers. Examples of conditions that the compositions of this invention are intended to prevent or alleviate include hypertension, cardiovascular disease, emphysema, and platelet aggregation. However, this composition also has a positive effect on many other conditions. The invention will be further described in connection with specific examples. Examples 1 and 2 are comparative examples in which the analysis of inositol phosphates in several commercially available bread and breakfast foods is carried out. Example 3
-7 illustrate the bread manufacturing method of this invention. Example 8 relates to the production of a vine cake made by baking flour with the addition of calcium salts of inositol phosphates. Example 9 relates to the production of a processed breakfast food after addition of sodium salts of inositol phosphates. Example 10 is D-myo-inositol-
1.2.6 - To illustrate the production of table salt by addition of the sodium salt of triphosphate. Example 11 illustrates the production of a drink by the addition of sodium salts of inositol triphosphates. example
No. 12 relates to the production of honey by addition of the sodium salt of D-myo-inositol-1.2.6-triphosphate. Example 13 relates to the production of thiocholate by addition of sodium salts of inositol phosphates. Example 14 shows that the aggregation of platelets caused by injection of cadmium in the blood of rabbits can be prevented by administration of a diet containing _IP3 . Example 15 shows the effect of IP ₃ on the cadmium content of various organs of cadmium-injected mice. Example 16 shows that IP ₃ prevents the increase in platelet aggregation caused by smoking. Examples 17 and 18 show that IP ₃ prevents or reduces the generation of free radicals. Examples 19-24 demonstrate the hydrolysis of phytic acid in various food sources. Examples 25-31 are
Figure 2 shows the preparation of IP ₃ and its separation into different isomers. Example 1 Analysis of inositol phosphates in several commercially available breads Three commercially available breads: one white bread and two crisp breads
was analyzed for inositol phosphate content using HPLC. The crispy breads are baked with whole rye flour and whole wheat flour, respectively. A 20g amount of bread was crushed and extracted with 1% hydrochloric acid for 2 hours with shaking. The suspension was centrifuged and the supernatant was collected. The supernatant was analyzed using well-defined inositol phosphates and quantified as mg of inositol phosphate per 100 g (solids content).

[Table] The results show that the amount of inositol triphosphate in commercial bread is small. Example 2 Analysis of inositol phosphates in breakfast foods Commercially available Corn Flakes
(trademark, manufactured by Kellotsugu Co., Ltd.) was analyzed for the content of inositol phosphates by HPLC.
The extraction method and analysis were similar to those described in Example 1. 19 mg IP ₂ and 2 mg IP ₃ were found per 100 g solids. IP ₄ , IP ₅ or IP ₆ could not be detected. The IP ₆ contained in the raw material was almost completely destroyed, and the amount of IP ₃ was very small. Example 3 Variation of fermentation time for crisp bread baked with rye Biologically acidified crisp bread was baked using rye flour (1% IP ₆ content). The dough composition is 54.6g of flour, 41.8g of water, 1.3g of salt (NaCl),
and 2.4 g of dough from the previous dough formulation. A sour dough consisting of 2.4 g of the above dough from the previous dough blend and 40% of the above flour and 85% of the above water was fermented in a first stage for 6 hours and then the other ingredients (flour, water and salt) were mixed. After mixing, the dough was allowed to rise in a second stage, followed by shaping and baking. Oven temperature 250
℃. Three types of bread were made using different second proofing times. The bread was crushed, extracted and analyzed as described in Example 1. The content of _IP3 versus fermentation time was determined as follows.

[Table] The results show that extended second fermentation time results in a decrease in IP ₃ content. Example 4 IP ₃ content in bread baked with wheat and oat flour Wheat and oat flour (IP ₆ content 0.9%) were combined to bake chemically acidified bread. The dough composition is 37.7% wheat flour and 17.7% oat flour.
%, water 39.5%, salt (NaCl 50% and calcium acetate 50%), sucrose 1%, and baker's yeast 2.8%
%. After mixing these ingredients, the dough was allowed to rise, then shaped and baked. Fermentation time is 90
minutes and the temperature was 37°C. The resulting bread was crushed, extracted and analyzed as described in Example 1. IP ₃ content dry bread 100
It was determined to be 120 mg/g. Example 5 IP ₃ content in bread baked with whole wheat flour White bread was baked with whole wheat flour (IP ₆ content 0.9%).
The dough composition is 55.9% flour, 38.4% water, 3.5% yeast,
Salt (NaCl) was 0.6%, and sucrose was 1.7%. After mixing the ingredients, the dough was allowed to rise and then the bread was formed. After additional fermentation time the bread was baked. Total fermentation time is 60 minutes, baking time is
The temperature was 175℃. The bread was crushed, extracted and analyzed as described in Example 1. IP ₃ content is 100g dry bread
It was 70mg per serving. Example 6 IP ₃ content in bread baked with rye flour with addition of _sodium phytate. %) was used to bake biologically acidified crispy bread. Fermentation time is 225
It was set as 1 minute. The bread was crushed, extracted and analyzed as described in Example 1. The amount of IP ₃ in the bread was 180 mg/100 g of dry bread. Example 7 IP ₃ content in bread baked on rye flour with addition of calcium magnesium phytate As described in Example 3 but with 100 g of dough
Using rye flour (IP ₆ content 1%) with the addition of 1.5 g of calcium magnesium phytate per
Biologically acidified crisp bread baked. Fermentation time was 225 minutes. The bread was crushed, extracted and analyzed as described in Example 1. The amount of IP ₃ in bread is per 100g of dry bread.
It was 250 mg. Example 8 IP ₃ content in a cake baked in wheat flour with addition of calcium salts of inositol phosphates A cake was baked in wheat flour (IP ₆ content 0.2%). The dough composition is 60.1% wheat flour, 35.7% water, and salt (NaCl).
0.6% and yeast 3.6%. After mixing the ingredients, the dough was allowed to rise. 0.2 g of a calcium salt of inositol phosphates containing 30% by weight IP ₃ was added to 100 ml of water per 100 g of dough and then the cake was formed. After additional rising time, the cake was baked in the oven. The total fermentation time was 75 minutes and the baking temperature was 225°C. The cake was crushed, extracted and analyzed as described in Example 1. The amount of IP ₃ was 60 mg/100 g of dry cake. Example 9 IP ₃ content in a processed breakfast food after addition of sodium salts of inositol phosphates 1000 g of commercially available corn flakes (trademark, manufactured by Kellogg) are mixed with 50% sucrose and 10% of sodium salts of inositol phosphates. (IP ₃
A hot (80°C) aqueous solution containing 30% content) was sprayed.
After drying, the breakfast food was crushed, extracted and analyzed as described in Example 1.
The content of IP ₃ was 30 mg/100 g of dry matter. Example 10 Table Salt Added with Sodium-Inositol Phosphate The sodium salt of D-myo-inositol-1.2.6-triphosphate was mixed with table salt to give a final concentration of IP ₃ of 200 ppm. Example 11 Drink with added sodium salt of inositol phosphates 20 ml of a 15% aqueous solution of the sodium salt of inositol phosphates (containing 40% IP ₃ )
of commercially available Coca-Cola (trademark), Seven-Up (trademark), and Orange Juice, respectively. Final concentration of _IP4 in drinks determined by HPLC: 190
mg/. Example 12 Honey with added sodium salts of inositol phosphates 5 ml of a 20% aqueous solution of the sodium salt of D-myo-inositol-1.2.6-triphosphate was added to 50 kg of commercial honey. The final concentration of inositol triphosphate was determined to be 20 mg/Kg by HPLC. Example 13 Thyokolate with IP ₃ To 1500 g of molten thiocholate, add 6 ml of a 10% aqueous solution of the sodium salt of inositol phosphates (containing 30% IP ₃ ), reduce the temperature and form the final thiocholate product. did.
The IP ₃ content in thiyocholate was 110 mg/kg thiyocholate as determined by HPLC. Example 14 Rabbits (New Zealand White, male) weighing 2-2.5 kg were used. These animals were fed inositol phosphate-free food for 10 days and then used in experiments. Two hours before the start of the experiment, myo-inositol-
1.2.6 - 50 mg of sodium triphosphate
Mixed into 5 g of feed for a group of 18 animals.
Another group of 12 animals did not receive inositol triphosphate.

[Table] Sample processing Two blood samples from each animal were run at 1200 rpm.
Centrifuge for 10 minutes and obtain plasma with platelets. Plasma with platelets from two samples was
Born [Journal of Physiology (J.
Physiol.), an aggregometer [Chronopar Corp model]
440], the response to the addition of ADP (adenosine diphosphate) was analyzed. Two samples were analyzed simultaneously at the same ADP concentration (1-20 micromolar) in two channels of the instrument. The principle of this test is that plasma with platelets becomes cloudy;
And it has low light transmittance. ADP
When added, the platelets aggregate and form clumps. This results in an increase in permeation, which is quantified by the device. Responses to ADP were measured in scale units with maximum aggregation of 80 scale units. To obtain the maximum sensitivity of this method to detect changes in platelet reactivity,
The amount of ADP should elicit a response of 5 to 30 scale units. This is generally achieved with 5 μM ADP, although lower or higher amounts (1-20 μM) were required in some animals. The results of this test are expressed as the maximum aggregation (scale units) of sample 2 minus the maximum aggregation of sample 1. The following results were obtained.

[Table] At the doses used in this experiment,
IP ₃ prevented the effect of Cd on platelet aggregation. Increased aggregation of platelets is considered to be one of the most important factors giving rise to cardiovascular diseases, such as arteriosclerosis, and the ability of IP ₃ to prevent cadmium-induced aggregation suggests that IP ₃ This has been shown to be very useful in preventing or alleviating such diseases. Example 15 Mice with a weight of 18-20 g at the beginning of the experiment were used. During the experiment and for at least 7 days prior to the experiment, mice were fed a semi-synthetic diet containing no inositol phosphates. mouse 2
divided into groups. They received saline and inositol triphosphate (IP ₃ ), respectively, by intraperitoneal injection daily for 9 days. IP ₃ dosage is 5.0
mg/day. The injection volume was 0.2ml. On the second day of the experiment, a second i.p.
~10 min later, all mice were injected intravenously with 2.5 microcuries of ¹⁰⁹ Cd as cadmium chloride in 50 μl saline. After the last intraperitoneal injection, the mice were sacrificed and several organs were removed and weighed. Radioactivity in various organs was measured by counting with a gamma counter. Radioactivity in the organs of animals treated with IP ₃ was compared to that of control animals treated with saline for the same time. In the results, the radioactivity in the organs of animals treated with IP ₃ was expressed as a % of the radioactivity found in the control. The results were as follows. Organ levels of mice treated with cadmium and IP ₃ are expressed as % control level (control = 100
%). Each group consists of 15 mice. The control group was
Treated with salt solution and Cd as above.

Table: The results show that IP ₃ causes a decrease in cadmium levels in all organs tested except liver and kidney. Cd is believed to be relatively safe at the latter site. Example 16 IP ₃ for platelet aggregation after smoking in humans
We investigated the effects of Young healthy male non-smokers were given two doses of capsules containing 50 mg of IP ₃ or 50 mg of a placebo. The _IP3 used was the calcium salt of D-myo-inositol-1.2.6-triphosphate. Both subjects and researchers were blinded to whether subjects were given IP ₃ or a placebo. Blood samples were obtained 2 hours after capsule ingestion. Next, the subject smoked two cigarettes in quick succession. A second blood sample was taken after this smoking. Platelet aggregation responses to ADP and collagen in these two samples were determined using substantially the same method as described in Example 14.
Results are expressed as the change in aggregation from pre-smoking samples to post-smoking samples. The + sign indicates that the aggregation became stronger after smoking.

[Table] In the placebo group, smoking increased aggregation;
This was most noticeable at low concentrations of flocculant. In all cases this effect was prevented by _IP3 . That is, IP ₃ prevented the increase in platelet aggregation induced by smoking. Example 17 48 mmol KH ₂ PO ₄ , 2 mmol sodium ascorbate, 0.1 mmol H ₂ O ₂ , 0.5 mlFe, and 1.7 mmol
The reaction mixture consisting of 37 mol deoxyribose
Incubate at ℃ for 1 hour. 1 mmol
Similar reaction mixtures containing EDTA or 1 mmol inositol triphosphate (IP ₃ ) were similarly incubated. D-Mio- as IP ₃
Inositol-1.2.6-triphosphate was used. After incubation, 1.65 ml of thiobarbituric acid and 1.65 ml of 2.8% trichloroacetic acid in 50 mmol NaOH were added to 2 ml of the reaction mixture. The mixture was heated to 100° C. for 20 minutes and the absorption at 532 nm was measured using water as a blank. The experiment was carried out using Fe ²⁺ [Fe(NH ₄ ) SO ₄ ] and Fe ³⁺
This was done using iron in the form of (FeCl ₃ ). The results were as follows. Free radical formation catalyzed by Fe ²⁺ and Fe ³⁺ in the presence of IP ₃ or EDTA is expressed as absorption at 532 nm. Group Fe ²⁺ Fe ³⁺ Control 0.76 0.79 EDTA 2.2 1.86 IP ₃ 0.46 0.43 These results show that the production of free radicals in the reaction mixture was reduced by 40% after the addition of IP ₃ . Addition of EDTA had the opposite effect. This strongly increased the production of free radicals. In this way, IP ₃ is a free radical ion −
It has been shown to reduce dependence generation. Example 18 Lipid peroxidation in lipid micelles was investigated. The next reaction mixture was incubated at 37°C for 2 hours. 0.4ml Clark Lubs buffer (PH5.5) 0.2ml Phospholipid liposomes 0.1ml 0.5-5mM IP ₃ or 0.1ml _H2O 0.1ml 1mM Fe2 ⁺ or _0.1mlH2O 0.1ml 4mM ^{Al3 +} or 0.1 ml _H2O 0.1 ml _H2O IP ₃ was D-myo-inositol-1.2.6-triphosphate. After incubation,
0.5ml thiobarbituric acid + 0.5ml 25% HCl was added and the mixture was heated at 100°C for 15 minutes. Lipid peroxidation was determined by adding 1 ml of Lubrol PX 1% (Sigma) and measuring absorbance at 532 nm. The results were as follows.

【table】

[Table] Fe ²⁺ induced lipid peroxidation (Group 1: 10).
Al ³⁺ itself did not cause peroxidation (group 2: 10)
However, the combination of Fe ²⁺ + Al ³⁺ caused much stronger peroxidation than Fe ²⁺ alone (1:3). Addition of IP ₃ completely prevented the interaction between Fe ²⁺ and Al ³⁺ (3:4). In Fe ²⁺ -only systems, IP ₃ induced a significant reduction in radical generation (1:5 ~
9). Example 19 Hydrolysis, extraction and IP of phytic acid in wheat ₃
Analysis of 100 g of crushed wheat seeds containing 1% myo-inositol hexaphosphate (IP ₆ )
ml of sodium acetate buffer PH5.2 at 35°C. After 30 minutes of incubation, the slurry was heated to -10°C to stop hydrolysis.
It was frozen at. 10 g of this frozen material was extracted with 100 ml of 0.4M HCl. The suspension was shaken for 1 hour and centrifuged. The supernatant was collected and neutralized to pH 7 with aqueous NaOH. A sample of the supernatant
Neutralized by HPLC. The analytical method was quantified using well-defined inositol phosphates. The IP ₃ content of the extract was 10 mg of inositol triphosphate. Example 20 Hydrolysis, extraction and IP ₃ analysis of phytic acid in white beans The same method as described in Example 19 was used. however,
100 g of white beans containing 1% myo-inositol hexaphosphate were incubated at 55° C. for 10 hours. 10 g of frozen material was extracted with 100 ml of 0.4M HCl. The suspension was shaken for 1 hour and then centrifuged. Collect the supernatant and dilute with NaOH aqueous solution.
Neutralized to PH7. A sample of the supernatant was analyzed by HPLC. The IP ₃ content of the extract was 5 mg. Example 21 Analysis of hydrolysis, extraction and IP ₃ of phytic acid in soybeans after addition of a microbially derived phytase source 300 g of soybeans were soaked overnight (IP ₆ content 1.4%), peeled and then boiled for 30 minutes. did. 1g of Rhizopus oligosporus
3 ml of water containing (NRRL 2710) were added and the mixture was incubated at 40° C. for 20 hours. 10 g of the mixture was extracted and analyzed by HPLC as described in Example 19. extract of
IP ₃ content was 160 mg. Example 22 Hydrolysis of phytic acid in white beans by crude wheat phytase, extraction and IP ₃ analysis Raw beans containing 1% myo-inositol hexaphosphate were suspended in 1000 ml of sodium acetate buffer pH 5.2. . 500 mg of crude wheat phytase (manufactured by Sigma Chemical Co.) was added. This mixture
Incubation was performed at 55°C with shaking. 12
After incubation for -10 hours, the slurry
The hydrolysis was stopped by freezing at ℃. 10 g of frozen material was extracted with 100 ml of 0.4M HCl.
The suspension was shaken for 1 hour and then centrifuged.
The supernatant was collected and neutralized to pH 7 with aqueous NaOH. A sample of the supernatant was analyzed by HPLC. The IP ₃ content of the extract was 40 mg. Example 23 IP ₃ content in white beans after addition of sodium phytate and hydrolysis 0.3 g of sodium phytate was added to 100 g of crushed white beans (IP ₆ 1%). This mixture was dissolved in 1000ml of sodium acetate buffer PH5.2 at 55%
Incubated at ℃. After 4 hours of incubation, the slurry was frozen to -10°C to stop hydrolysis. 10 g of frozen material was extracted and analyzed as described in Example 19. extract of
IP ₃ content was 15 mg. Example 24 1% inositol hexaphosphate (IP ₆ )
1.0 kg of rice bran containing 1.0 kg of rice bran was suspended in 10% sodium acetate buffer (PH5) at 25°C. After 4 hours and 50% of inorganic phosphorus has been liberated, the slurry is
of 2M HCl. The suspension was shaken for 1 hour and then centrifuged. Supernatant Ca
Neutralized to pH 7 with an aqueous solution of (OH) ₂ . 5 was added to obtain a precipitate. Calcium salts of various inositol phosphate compositions were centrifuged, dried, and recrystallized. 20 mg of recrystallized calcium salt was brought into acid form by addition of dilute hydrochloric acid and analyzed by HPLC. This composition contained 20% inositol triphosphate. The remainder were other inositol phosphates. Example 25 Hydrolysis of sodium phytate by wheat phytase and fractionation of a mixture of inositol phosphates 1.6 g of sodium phytate (derived from corn, manufactured by Sigma Chemical) was dissolved in 650 ml of sodium acetate buffer (PH 5.2). . 2.7g of wheat phytase (EC3.1.3.26, 0.015U/mg, manufactured by Sigma Chemical) was added and the mixture was incubated at 38°C. Dephosphorylation was followed by measuring the inorganic phosphorus released. After 3 hours, when 50% of the inorganic phosphorus was liberated, the hydrolysis was stopped by adding 30 ml of ammonia to bring the pH to 12. A liquid mixture containing inositol phosphates was obtained. Pass 350 ml of the mixture through an ion exchange column (Dowex 1, chloride type, 25 mm x 250 mm).
It was then eluted with a linear gradient of hydrochloric acid (0-0.7N). An aliquot of the elution fraction was completely hydrolyzed to determine the phosphorus and inositol content. Multiple peaks corresponded to different inositol phosphates. That is, a peak with a phosphorus to inositol ratio of 3:1 is composed of inositol triphosphate, etc. Two fractions were obtained with a 3:1 ratio of phosphorus to inositol. Example 26 Fractionation of inositol triphosphate 100 ml of the first fraction obtained in Example 25 with a phosphorus/inositol ratio of 3:1 was neutralized and
After adding a 10% excess of 0.1 M barium acetate solution, the barium salt was precipitated. 600 mg of precipitated salt was dissolved in 50 ml of dilute hydrochloric acid. This solution was separated on an ion exchange column (Dowex 1, chloride form, 25 mm x 2500 mm) using dilute hydrochloric acid as the eluent. An aliquot of the eluted fraction was analyzed for phosphorus. Three peaks consisting of isomers of inositol triphosphate could be observed. Example 27 Structural determination of isomers of inositol triphosphate by NMR The three peaks obtained in Example 26 were combined with H-
Analyzed by NMR. This data indicates that these peaks are respectively myo-inositol-1.2.6-
Triphosphate, myo-inositol-1.2.3
-Triphosphate and myo-inositol-
1.3.4- indicates that it consists of triphosphate. The second fraction obtained in Example 25 with a phosphorus/inositol ratio of 3:1 was analyzed by H-NMR. This data indicates that this fraction consists of myo-inositol-1.2.5-triphosphate. Example 28 Determination of optical isomers of inositol triphosphate 20 mg of the compounds determined by H-NMR of Example 27 to be myo-inositol-1.2.6-triphosphate and myo-inositol-1.3.4-triphosphate were further added. , chromatographed on an acetylated cellulose-based chiral column (20 mm x 300 mm, Merck) using a mixture of ethanol and water as eluent. Fractions were analyzed by polarimetry. Each compound has one optical isomer, D-myo-inositol-1.2.6
-triphosphate, and L-myo-inositol-1.3.4-triphosphate. Example 29 Hydrolysis of sodium phytate by baker's yeast and fractionation of a mixture of inositol phosphates 0.7 g of sodium phytate (derived from corn, manufactured by Sigma Chemical) was dissolved in 600 ml of sodium acetate buffer (PH 4.6). . Ja¨stblaget, baker's yeast from Sweden (28% dry matter, 2% nitrogen content, 0.4% phosphorus content) 50 g
was added with stirring, and incubation was carried out at 45°C. Dephosphorylation was followed by measuring liberated inorganic phosphorus. After 7 hours, when 50% of the inorganic phosphorus was liberated, the hydrolysis was stopped by adding 30 ml of ammonia to bring the pH to 12. The suspension was centrifuged and the supernatant was collected. 400 ml of supernatant was passed through an ion exchange column (Dowex 1, chloride form, 25 mm x 250 mm) and a linear gradient of hydrochloric acid (0 to 0.7 N HCl).
It was eluted by To determine the content of phosphorus and inositol,
An aliquot of the elution fraction was completely hydrolyzed. Each peak corresponded to a different inositol phosphate. That is, the ratio of phosphorus to inositol is 3:1.
The peak that is composed of inositol triphosphate...etc. Example 30 Structural Determination of Isomers of Inositol Triphosphate The fractions obtained in Example 29 with a phosphorus/inositol ratio of 3:1 were neutralized and evaporated,
After this, H-NMR analysis was performed. The data show that this peak consists of myo-inositol-1.2.6-triosphate. Example 31 Determination of optical isomers of myo-inositol triphosphate The same method as described in Example 28 was used. However, 10 mg of the compound determined by NMR in Example 30 was analyzed. As can be seen, this compound consists of one optical isomer, D-myo-inositol-1.2.6-triphosphate. In order to better understand this invention, the formulas of several IP ₃ isomers of this invention are provided. _IP6 , _IP5 ,
Formulas for IP ₄ and IP ₂ are also listed. Lower phosphate esters of myo-inositol are named according to where the phosphate group is located on the inositol ring, trying to give the lowest possible position number. The carbon atom bearing the axial phosphate group always has position number 2. The structural formulas below are simplified to the acid form.

【table】

[Table] In the above structural formula, =-O-PO ₃ H ₂ .

Claims (1)

[Claims] 1. A method for producing a food composition, which comprises first adding to a food composition substantially free of inositol triphosphate (IP ₃ ) a final concentration of 5 mg or more of IP ₃ per 100 g of the composition. a method comprising adding a source of _IP3 in an amount sufficient to provide a. 2. The method of claim 1, wherein the _IP3 source is _IP3 . 3. The method according to claim 1, wherein the source of _IP3 is inositol hexaphosphate ( _IP6 ), inositol pentaphosphate ( _IP5 ) and/or inositol tetraphosphate ( _IP4 ) in the presence of phytase. Method. 4. The IP ₃ source is the following IP ₃ isomers: D-myo-inositol-1,2,6-triphosphate, D-myo-inositol-1,2,5-triphosphate, myo-inositol-1,2, 3
-Triphosphate, L-myo-inositol-
1,3,4-triphosphate, or D-myo-
The method according to claim 1, wherein at least one inositol-1,4,5-triphosphate is used. 5. Establishing the content of phytase and the content of inositol phosphate selected from the group consisting of IP ₄ , IP ₅ and IP ₆ in the materials making up the composition, and in at least one step of the manufacturing process,
2. The method according to claim 1, wherein the time and temperature of the treatment and the pH value are controlled so that the incubation can be carried out so as to obtain an IP ₃ content of 20 mg or more per 100 g of the composition. 6. The method of claim 1, wherein the composition is a solid and the content of IP ₃ is greater than or equal to 20 mg per 100 g of dry composition. 7. The method of claim 1, wherein said composition is a cereal-based material. 8. The method of claim 7, wherein said composition is selected from the group consisting of breakfast foods, cakes and biscuits. 9. The method of claim 1, wherein said composition is selected from the group consisting of confectionery, tyokolate, and chewing gum. 10. The method of claim 1, wherein said composition is a vegetable. 11. The method of claim 6, wherein the composition is bread and the content of _IP3 is 50 mg or more per 100 g of dry bread. 12. The method of claim 1, wherein said composition is selected from the group consisting of beverages and soups. 13. The method of claim 5, wherein the incubation is carried out separately and the resulting IP ₃ -containing product is added to the composition to control the desired IP ₃ content. 14. The method according to claim 3, wherein the phytase is added in the form of yeast. 15 Temperature range at incubation stage is 20
6. The method according to claim 5, wherein the temperature is 0.degree. C. to 70.degree. C. and the pH value is in the range 4 to 8. 16 IP ₃ concentration is 20 mg to 500 per 100 g of dry composition
The method according to claim 1, wherein the method is mg. 17 IP ₃ concentration is 50 mg to 500 per 100 g of dry composition
The method according to claim 1, wherein the method is mg. 18 The composition is in liquid form and has an IP ₃
A method according to claim 1, wherein the concentration is from 5 mg to 300 mg per 100 g of said liquid composition. 19. A method of preparing a food composition from a composition initially containing less than about 10 mg of IP ₃ per 100 g of the composition, the content of said IP ₃ being reduced by the addition of IP ₃ or a source of IP ₃ ; ₆ , IP ₅ and IP ₄ by conversion by enzymatic treatment to increase the amount of inositol phosphate to 20 mg or more per 100 g of the composition. 20. The method of claim 19, wherein said composition is selected from the group consisting of cereal-based ingredients, confectionery, tyokolate, chewing gum, vegetables, fruits, beverages, and soups. 21. The method according to claim 19, wherein the content of IP ₃ is increased to 30 mg or more and preferably 50 to 500 mg per 100 g of the composition. 22 A method for producing a food composition, which comprises determining the amount of phytase and IP ₄ in the materials constituting the composition;
Establish an amount of inositol phosphate selected from the group consisting of IP ₅ and IP ₆ , and a sufficient amount of IP ₃
characterized by controlling the processing time and temperature as well as the PH value to allow incubation so that the degree of hydrolysis of IP ₄ , IP ₅ and IP ₆ is 40-70% to obtain Method. 23 The concentration of IP ₃ is 5 mg or more per 100 g of dry composition and IP ₃ is added to the composition and/or phytase and IP ₄ , IP ₅ and IP ₆
A food composition containing IP ₃ , characterized in that it is produced by the addition of an inositol phosphate selected from the group consisting of: 24. The composition of claim 23, wherein the food composition is a non-grain food and has an IP ₃ content of 5 mg or more per 100 g of the composition. 25. A composition according to claim 24, wherein the content of _IP3 is 10 mg per 100 g of the composition and preferably ranges from 20 to 500 mg. 26. The composition of claim 24, wherein said composition is selected from the group consisting of confectionery, tyokolate, chewing gum, vegetables, fruits, drinks and soups. 27 The above IP ₃ is the following isomers, namely D-myo-inositol-1,2,6-triphosphate, D-myo-inositol-1,2,5-triphosphate, myo-inositol-1,2,3
-Triphosphate, L-myo-inositol-
1,3,4-triphosphate, or D-myo-
25. The composition according to claim 24, which is at least one inositol-1,4,5-triphosphate. 28 The food composition is a bread food, the concentration of IP ₃ is 20 mg or more per 100 g of the dry composition, and the degree of hydrolysis of naturally contained IP ₆ and/or added IP ₆ is 40 to 70 % of the composition of claim 23. 29. A composition according to claim 28, wherein the concentration of _IP3 is greater than or equal to 30 mg and preferably from 50 to 500 mg per 100 g of dry composition. 30. The composition of claim 28, wherein the bread food is selected from the group consisting of bread, breakfast food, biscuits, and cakes. 31. The composition of claim 28, wherein said _IP3 is substantially D-myo-inositol-1,2,6-triphosphate. 32 said IP ₃ is a hydrolysis product derived from an inositol phosphate selected from IP ₄ , IP ₅ and IP ₆ , wherein the distribution curve indicating the content of different inositol phosphates has a maximum for IP ₃ ; The composition according to any one of claims 24 to 28. 33. The composition according to claim 32, wherein the content of IP ₃ is greater than 40% by weight of the total amount of inositol phosphates and 30-85% by weight of the remaining inositol phosphates consists of IP ₂ and IP ₄ . . 34. The composition of claim 24 in a concentrated form comprising IP ₃ in a food-acceptable filler.